Host prenylation promotes membrane association of the Salmonella effector GtgE during infection

The Rab32 antimicrobial pathway restricts Salmonella Typhi replication in murine macrophages. The broad-host pathogen Salmonella Typhimurium has evolved a solution to disrupt this pathway via effector proteins, including GtgE. The presence of a C-terminal CaaX motif in GtgE prompted the hypothesis that the effector was prenylated by the host. Here, we show that the CaaX motif enhances GtgE membrane association and intracellular targeting. Substitution of the C-terminal cysteine (C225S) caused diffused cytosolic distribution and loss of membrane enrichment. Inhibition of host isoprenoid biosynthesis or geranylgeranyl transferase I activity also disrupted GtgE localization. During infection, translocated GtgE was enriched in membrane fractions and accumulated around intracellular Salmonella in a pattern consistent with association to the Salmonella -containing vacuole, whereas the C225S variant showed reduced membrane association. These findings identify host prenylation as a mechanism that may direct GtgE to intracellular membranes where Rab32 is accessible for cleavage.